The present invention relates generally to the field of immunology.
Induction of immunity to pathogens, toxins, and peptides expressed by tumor cells, requires the coordinated participation of the innate and adaptive immune systems. An early step is Ag internalization by APCs of the innate immune system, notably by dendritic cells (DCs), the most potent APC type, and the one best able to present Ag to naïve T cells (Trombetta and Mellman, 2005). Internalized Ag is processed through the endosomal/lysosomal path. Processed peptides, bound to MHC molecules, are then delivered to the cell surface. Those T cells with appropriate receptors respond to such peptides provided co-stimulatory molecules are expressed by the DC. A second signal is often required to drive DC maturation and efficient co-stimulatory molecule expression. Ag activates B cells bearing appropriate surface immunoglobulin directly to produce IgM. CD4+ T cells, having responded to processed Ag, induce immunoglobulin class-switching from IgM to IgG.
Limited uptake of soluble antigenic peptide by DCs constrains subsequent Ag processing and presentation. Immune responses increase when Ag uptake is facilitated. IgG-immune complexes (ICs) bind to FcγRs expressed on DCs and this is followed by internalization of ICs with their captured Ags. Thus, stronger Ab responses may occur when soluble Ag is complexed to IgG, than when Ag alone is administered (Wemersson et al., 1999). ICs in antibody excess can be more effective at Ag presentation than ICs at equivalence or in Ag excess (Manca et al., 1991). IC driven, FcγR-mediated, Ag internalization favors DC maturation and hence expression by them of costimulatory molecules (Regnault et al., 1999). Other means to target Ag to Fcγ receptors on APCs have been employed in order to elicit strong immune responses against otherwise weak immunogens. Early studies, that documented the potential of this approach, employed Ag-containing anti-FcγR monoclonal antibodies as a means to facilitate delivery of Ag to APCs and hence increase Ag-specific T cell responses and Ag-specific humoral responses (Snider et al., 1990; Heijnen et al., 1996; Gosselin et al., 1992; Keler et al., 2000). Modification of Ig by introduction of epitopes within the CDR region (i.e., antigenized Ig) may also enhance immune responses compared to Ag alone (Zaghouani et al., 1993, Brummeanu et al., 1996).
Immune complexes (IC) exhibit diverse biological activities; some that contribute to disease whereas others ameliorate disease. Deposition of IgG containing IC on tissue surfaces, as for example in glomeruli, can contribute to the pathogenesis of antibody-mediated autoimmune diseases. On the other hand, IC can favorably modulate T- and B-cell activation pathways via binding to Fc receptors expressed on immunocytes. Aggregated IgG (AIG) shares some features and biological activities with IC. Both modulate T-cell suppressor function (Antel et al., 1981; Durandy et al., 1981), cytokine synthesis, IgG secretion, and lymphocyte proliferation (Berger et al., 1997; Wiesenhutter et al., 1984; Ptak et al., 2000).
Monomeric IgG, or the Fc fragment thereof, can ameliorate disease progression in animal models of autoimmune disease (Miyagi et al., 1997; Gomez-Guerrero et al., 2000). Monomeric IgG can be used therapeutically, usually in massive doses, to treat antibody-mediated diseases in man. The protective effect in antibody-mediated diseases may be achieved in part through blockade of FcγRs such that binding of IC to them is impeded (Clynes et al., 1998). IgG administration also favorably affects the course of T-cell mediated autoimmune diseases such as multiple sclerosis (Fazekas et al. 1997; Sorensen et al., 1998; Achiron et al., 1998). Here the basis for benefit is poorly understood though it is postulated to involve the increased production of anti-inflammatory cytokines initiated by binding of IV IgG, or complexes derived therefrom, to FcγR. In both antibody and T-cell mediated processes the mechanisms and consequences of FcγR engagement are fundamental to the understanding and treatment of autoimmune diseases.
Aggregated IgG has been proposed as a treatment for autoimmune diseases of humans. The use of aggregated IgG has been studied as a treatment for multiple sclerosis and other autoimmune diseases. However, aggregated IgG has major limitations. IgG is commonly aggregated by exposure to heat; the resultant aggregates are bound together in a random fashion limiting reproducibility from one preparation to the next. Preparations contain a heterogeneous collection of aggregates of varying size in diverse conformations.
U.S. Pat. Nos. 5,714,147 and 5,455,165 disclose some hybrid immunoglobulin molecules and the expression vectors encoding them. These chimeric molecules can improve the circulating plasma half-life of ligand binding molecules, and can comprise a lymphocyte homing receptor fused to an immunoglobulin constant region. Homo or hetero-dimers or tetramer hybrid immunoglobulins containing predominantly the heavy and light constant regions of immunoglobulin have been used. U.S. Pat. No. 6,046,310 discloses FAS ligand fusion proteins comprising a polypeptide capable of specifically binding an antigen or cell surface marker for use in treatment of autoimmune disorders. The fusion protein preferably comprises IgG2 or IgG4 isotype, and may comprise antibodies with one or more domains, such as the CH2, CH1 or hinge deleted. Majeau et al. (1994) discusses Ig fusion proteins used for the inhibition of T cell responses. These fusion proteins comprise IgG1 and LFA-3. Eilat et al. (1992) disclose a soluble chimeric Ig heterodimer produced by fusing TCR chains to the hinge region, CH2, and CH3 domains of human IgG1.
Immunoglobulin fusion proteins can be employed to express proteins in mammalian and insect cells (Ashkenazi, et al., 1997). Fusion protein platforms can permit the introduction of additional functions, for example, inclusion of the amino-terminal CD8α domain may result in the co-ligation of FcR on lymphocytes to MHC I on antigen presenting cells (Alcover, et al., 1993; Meyerson, et al., 1996).
Other Ig proteins and variants have also been studied for their therapeutic effect on autoimmune diseases, including a recombinant polymeric IgG that mimics the complement activity of IgM (Smith and Morrison, 1994) where the polymeric IgG is formed by the polymerization of H2L2 subunits. Greenwood et al. (1993) discusses therapeutic potency relative to the structural motifs involving the human IgG antibodies, IgG1, IgG3, and IgG4. U.S. Pat. No. 5,998,166 discloses human FcγR-III variants, which can be used in the therapy or diagnosis of autoimmune diseases. U.S. Pat. No. 5,830,731 discloses novel expression vectors in which cell surface antigens cloned according to that invention appear to have diagnostic and therapeutic utility in immune-mediated infections. Cell surface antigens that are used to regulate lymphocyte activation, appear to achieve antigen aggregation in vitro by incubating lymphocytes with immobilized ligands or antibodies or their fragments (WO9942077). However, the aggregated IgG and Fc aggregates have limited reproducibility, containing a random and heterogeneous mixture of protein thereby limiting their effectiveness as therapeutic agents. Other problems include a lack of an ability to target a number of cell types with a single agent and size limitations.